Physiological Importance of Pectin Modifying Genes During Rice Pollen Development.

Although cell wall dynamics, particularly modification of homogalacturonan (HGA, a major component of pectin) during pollen tube growth, have been extensively studied in dicot plants, little is known about how modification of the pollen tube cell wall regulates growth in monocot plants. In this study, we assessed the role of HGA modification during elongation of the rice pollen tube by adding a pectin methylesterase (PME) enzyme or a PME-inhibiting catechin extract (Polyphenon 60) to in vitro germination medium. Both treatments led to a severe decrease in the pollen germination rate and elongation. Furthermore, using monoclonal antibodies toward methyl-esterified and de-esterified HGA epitopes, it was found that exogenous treatment of PME and Polyphenon 60 resulted in the disruption of the distribution patterns of low- and high-methylesterified pectins upon pollen germination and during pollen tube elongation. Eleven PMEs and 13 PME inhibitors (PMEIs) were identified by publicly available transcriptome datasets and their specific expression was validated by qRT-PCR. Enzyme activity assays and subcellular localization using a heterologous expression system in tobacco leaves demonstrated that some of the pollen-specific PMEs and PMEIs possessed distinct enzymatic activities and targeted either the cell wall or other compartments. Taken together, our findings are the first line of evidence showing the essentiality of HGA methyl-esterification status during the germination and elongation of pollen tubes in rice, which is primarily governed by the fine-tuning of PME and PMEI activities.

Integrative analysis of pectin methylesterase (PME) and PME inhibitors in tomato (Solanum lycopersicum): Identification, tissue-specific expression, and biochemical characterization.

Although previous studies have demonstrated that the degree of demethylesterification of pectin polysaccharides is modulated during tomato fruit ripening, its involvement in vegetative organ development has been seldom investigated. As a first step in understanding the importance of pectin modification during vegetative stages, we used chemical, biochemical, and molecular approaches to analyze PMEs and PMEIs in tomato plants. We found that tomato cell walls isolated from vegetative tissues as well as the fruit contain substantial quantities of pectin, and different degrees of methylesterification were evident in different tissues. Our chemical study was further substantiated by immunolocalization analysis, which showed that selective removal of pectin-bound methyl groups is required for proper organ development and growth. In the tomato genome, there exists 79 PMEs and 48 PMEIs with temporally and spatially regulated expression. As a case study, we showed that two tomato PMEIs (SolycPMEI13 and SolycPMEI14) exhibited PMEI activities. This is the first report regarding the genome-wide identification and expression profiling of PME/PMEIs in tomato and the first chemical evidence of the differential degrees of pectin methylesterification in vegetative and reproductive tissues. Taken together, our findings provide an important tool to unravel the molecular and physiological functions of tomato PME and PMEI in further study.

Rice pectin methylesterase inhibitor28 (OsPMEI28) encodes a functional PMEI and its overexpression results in a dwarf phenotype through increased pectin methylesterification levels.

Pectin methylesterases (PMEs, EC 3.1.1.11) belonging to carbohydrate esterase family 8 cleave the ester bond between a galacturonic acid and an methyl group and the resulting change in methylesterification level plays an important role during the growth and development of plants. Optimal pectin methylesterification status in each cell type is determined by the balance between PME activity and post-translational PME inhibition by PME inhibitors (PMEIs). Rice contains 49 PMEIs and none of them are functionally characterized. Genomic sequence analysis led to the identification of rice PMEI28 (OsPMEI28). Recombinant OsPMEI28 exhibited inhibitory activity against commercial PME protein with the highest activities detected at pH 8.5. Overexpression of OsPMEI28 in rice resulted in an increased level of cell wall bound methylester groups and differential changes in the composition of cell wall neutral monosaccharides and lignin content in culm tissues. Consequently, transgenic plants overexpressing OsPMEI28 exhibited dwarf phenotypes and reduced culm diameter. Our data indicate that OsPMEI28 functions as a critical structural modulator by regulating the degree of pectin methylesterification and that an impaired status of pectin methylesterification affects physiochemical properties of the cell wall components and causes abnormal cell extensibility in rice culm tissues.

Genome-wide identification and expression analysis of rice pectin methylesterases: Implication of functional roles of pectin modification in rice physiology.

Pectin, which is enriched in primary cell walls and middle lamellae, is an essential polysaccharide in all higher plants. Homogalacturonans (HGA), a major form of pectin, are synthesized and methylesterified by enzymes localized in the Golgi apparatus and transported into the cell wall. Depending on cell type, the degree and pattern of pectin methylesterification are strictly regulated by cell wall-localized pectin methylesterases (PMEs). Despite its importance in plant development and growth, little is known about the physiological functions of pectin in rice, which contains 43 different types of PME. The presence of pectin in rice cell walls has been substantiated by uronic acid quantification and immunodetection of JIM7 monoclonal antibodies. We performed PME activity assays with cell wall proteins isolated from different rice tissues. In accordance with data from Arabidopsis, the highest activity was observed in germinating tissues, young culm, and spikelets, where cells are actively elongating. Transcriptional profiling of OsPMEs by real-time PCR and meta-analysis indicates that PMEs exhibit spatial- and stress-specific expression patterns during rice development. Based on in silico analysis, we identified subcellular compartments, isoelectric point, and cleavage sites of OsPMEs. Our findings provide an important tool for further studies seeking to unravel the functional importance of pectin modification during plant growth and abiotic and biotic responses of grass plants.